IRS-2 is a serpin with activity against chymotrypsin-like, rather than trypsin-like, serine proteases. (A) Proteases targeted by IRS-2. α-chymotrypsin, chymase, cathepsin G, trypsin, and thrombin are inhibited by IRS-2. The amount of enzyme used is stated in Table 1. The mean remaining enzymatic activity in the presence of various concentrations of IRS-2 is represented, while the error bars represent the SEM in triplicate assays. (B) Inhibition data are normalized by plotting the remaining enzymatic activity (y-axis) against the IRS-2/enzyme ratio (x-axis). (C) Inhibition of purified mMCP-4 (5nM) or present in the suspension of activated mouse PCMCs by IRS-2. The mean remaining enzymatic activity in the presence of various concentrations of IRS-2 is presented (± SEM). (D) Western blot analysis showing covalent complex formation between IRS-2 and mMCP-4 produced by PCMCs. Notably, enzymatic activity and complexes between mMCP-4 and IRS-2 were predominantly cell associated, rather than being present in cell-free supernatants. Cells activated by ionomycin were used as positive control (+), and untreated cells were used as negative control (−). All samples with IRS-2 were activated by ionomycin.